Immunoglobulins D and M mediate signals that are qualitatively different in B cells with an immature phenotype.

Abstract

The CH family of murine B-cell lymphomas includes several members that are sensitive to growth inhibition when their membrane IgM (mIgM) receptors are crosslinked by anti-p chain, anti-c chain, or anti-idiotypic antibodies. These lymphomas are IgM+, Ia+, and IgD and resemble neonatal B cells in terms of their exquisite sensitivity to anti-IgM-mediated negative signaling as a model for tolerance induction. In this report, we describe the properties of one of these lymphomas, CH33, which had been transfected with a construct containing an allotypically different 8 chain constant region and the heavy chain variable region fragment from S107 The CH33 cell line has been used as a model for immature B cells because it expresses mIgM but only low levels of mIgD (6). These cells, like other immature B-cell lymphomas (6), are blocked at the G1/S interface when exposed to low concentrations of anti-IgM antibodies. We have used a CH33 cell line transfected with an exogenous, allotypically different, 8 chain gene construct containing a heavy chain variable region (VH) of known specificity to study IgD-mediated signaling. In addition, we have examined signaling via IgD in selected spontaneous derivatives of CH33 showing higher expression of endogenous mIgD than in the parental line. Our data suggest that mIgD and mIgM deliver qualitatively different signals to immature B cells and that IgD alone is not sufficient to change the functional phenotype of these cells. MATERIALS AND METHODSCell Lines and Culture Conditions. The characteristics of CH33 cells (donated by Geoffrey Haughton, University of North Carolina, Chapel Hill) have been described (6, 7). Briefly, the lymphoma was induced by a hyperimmunization protocol in B10.H-2aH-4bp/Wts mice and adapted to continuous growth in RPMI 1640 medium (GIBCO) supplemented with 5% (vol/vol) fetal bovine serum (HyClone, Logan, UT), 50 AuM 2-mercaptoethanol, 2 mM L-glutamine, 1 mM sodium pyruvate, nonessential amino acids, 10 mM Hepes, and 100 units of penicillin and 100 ug of streptomycin per ml in humidified 7% C02/93% air at 370C. ECH408

Inhibition of c-myc Expression Induces Apoptosis of WEHI 231 Murine B Cells

“…These cells do not cease proliferation, and apoptosis does not occur (54). Similar results were obtained by Ales-Martinez and coworkers with another B-cell line, CH33, also transfected to express IgD (2). Furthermore, mutants of WEHI 231 resistant to anti-IgM-induced apoptosis display only a transient increase in c-myc RNA levels following anti-Ig treatment (29), whereas treatment of WEHI 231 cells with transforming growth factor ␤ (TGF-␤) induces a drop in levels of c-myc expression and subsequently apoptosis (56).…”

Role of Phosphatidylinositol 3-Kinase in Anti-IgM- and Anti-IgD-Induced Apoptosis in B Cell Lymphomas

“…Our laboratory has investigated mechanisms by which the B cell Ag receptor (BCR) 4 induces apoptosis in several B cell lines as a model for tolerance. Others and we have shown that anti-IgM (anti-heavy chain, anti-) and not anti-IgD (anti-␦ heavy chain, anti-␦) induces growth arrest and apoptosis (2)(3)(4)(5)(6). Furthermore, we have also established that anti-, but not anti-␦, stimulates an increase in expression of the cyclin-dependent kinase (CDK) inhibitor, p27 Kip1 , which is concomitant with a net loss in c-Myc protein expression (5,7,8).…”

“…In contrast to anti-, anti-IgD (anti-␦) treatment causes neither growth arrest nor apoptosis in the ECH408, as well as other surface IgD (sIgD) expressing lines (2,3,17,25). The results presented in Table I Kip1 protein expression in other B cell lymphoma lines (7,8,38,39).…”

“…The murine B cell lymphoma line, ECH408, was derived from the CH33 clone transfected with a murine ␦-chain gene construct (2). The ECH408 line has been defined as functionally immature based on its sensitivity to anti--induced growth arrest and apoptosis and its high /␦ high surface Ig expression profile (4,25,26).…”

Association between B-lymphocyte membrane immunoglobulin and multiple members of the Src family of protein tyrosine kinases.

“…Following exposure for fluorography, individual bands were excised for phosphoamino acid analysis (26) and phosphotryptic peptide mapping (33 (1,6,44). The in vitro phosphorylation of proteins associated with membrane IgM and IgD was next compared (Fig.…”